Gene Expression Profile of Dendritic Cell-Tumor Cell Hybrids Determined by Microarrays and Its Implications for Cancer Immunotherapy.

Jens Dannull,Chunrui Tan,Scott Pruitt,Christine Farrell,Cynthia Wang,Smita K Nair,Walter T Lee

doi:10.1155/2015/789136

Jens Dannull, Chunrui Tan + Show 5 more

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https://doi.org/10.1155/2015/789136

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Abstract

Background. Dendritic cell- (DC-) tumor fusion cells stimulate effective in vivo antitumor responses. However, therapeutic approaches are dependent upon the coadministration of exogenous 3rd signals. The purpose of this study was to determine the mechanisms for inadequate 3rd signaling by electrofused DC-tumor cell hybrids. Methods. Murine melanoma cells were fused with DCs derived from C57BL/6 mice. Quantitative real-time PCR (qPCR) was used to determine relative changes in Th (T helper) 1 and Th2 cytokine gene expression. In addition, changes in gene expression of fusion cells were determined by microarray. Last, cytokine secretion by fusion cells upon inhibition of signaling pathways was analyzed by ELISA. Results. qPCR analyses revealed that fusion cells exhibited a downregulation of Th1 associated cytokines IL-12 and IL-15 and an upregulation of the Th2 cytokine IL-4. Microarray studies further showed that the expression of chemokines, costimulatory molecules, and matrix-metalloproteinases was deregulated in fusion cells. Lastly, inhibitor studies demonstrate that inhibition of the PI3K/Akt/mTOR signaling pathway could restore the secretion of bioactive IL-12p70 by fusion cells. Conclusion. Our results suggest that combining fusion cell-based vaccination with administration of inhibitors of the PI3K/Akt/mTOR signaling pathway may enhance antitumor responses in patients.

Highlights

Dendritic cells (DCs) have been identified as a key component in manipulating and stimulating the immune system [1]
We show that production of the Th1 skewing cytokine IL-12 was dramatically downregulated in Dendritic cell- (DC-)tumor fusion cells
Fusion cells were purified by FACS and RNA isolated from hybrid cells was analyzed by quantitative real-time PCR for expression levels of mRNAs encoding the Th1 cytokines IFN-γ, TNF-α, IL-2, IL-12p40, IL-15, and IL18 or the Th2 cytokines IL-4, IL-10, IL-13, and IL-25

Summary

Introduction

Dendritic cells (DCs) have been identified as a key component in manipulating and stimulating the immune system [1]. Activated DCs are potent antigen presenting cells that express both major histocompatibility complex (MHC) class I and II molecules (Signal 1) and costimulatory molecules (Signal 2) and secrete immune modulating cytokines (Signal 3) resulting in activation of T lymphocytes [2]. Depending on the cytokine environment, DCs may elicit either a Th (T helper) 1 or Th2 CD4 T-cell response. Changes in gene expression of fusion cells were determined by microarray. Cytokine secretion by fusion cells upon inhibition of signaling pathways was analyzed by ELISA. Inhibitor studies demonstrate that inhibition of the PI3K/Akt/mTOR signaling pathway could restore the secretion of bioactive IL-12p70 by fusion cells. Our results suggest that combining fusion cell-based vaccination with administration of inhibitors of the PI3K/Akt/mTOR signaling pathway may enhance antitumor responses in patients

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